Effect of an altitudinal gradient on the morphology, molecular identification and distribution of Rhipicephalus linnaei in Veracruz, Mexico.

Studies of morphological and genetic variation in vector populations across environmental gradients can help researchers to estimate species' responses to climate change scenarios and the potential risk of disease-causing pathogen expansion, which impacts negatively on human health. In this study, we analysed the effect of altitudinal gradients on the phenotypic response of the hard tick of medical and veterinary importance, Rhipicephalus sanguineus sensu lato (s.l.). Specimens of R. sanguineus s.l. were collected from host animals in one of Mexico's regions with high climatic heterogeneity (Veracruz), and geometric morphometric theory was employed to assess the response of three morphological characters to the altitudinal gradient. Additionally, genetic similarity data were provided, and ecological niche models were used to project the climatic distribution in the region. Our results demonstrate that the shape and size of ticks respond to altitude. Molecular identification indicate that all analysed samples correspond to the tropical lineage recently named Rhipicephalus linnaei. According to ecological niche models, the mean annual temperature contributes significantly to the spatial distribution of this tick species, with areas of higher suitability in the mountainous region. These changes in morphological structure and the presence of ticks at higher altitudinal gradients suggest that R. linnaei has a high potential for adaptation. Due to the variability of ecosystems in the state of Veracruz, our results could be valuable in assessing the response of this tick in a changing environment, aiding in predicting future scenarios in the distribution and abundance of this species.

Gestational monitoring in buffalo females: detection of Neospora caninum by PCR.

This study establishes the serological frequency against Neospora caninum on day zero and the presence of N. caninum DNA surveyed throughout the gestation of Bubalus bubalis females in a bovine buffalo system in the central zone of the state of Veracruz, Mexico. Blood samples were taken from 11 females in 6 different sampling periods and analyzed for N. caninum antibodies detection on day zero. DNA detection by PCR was performed on all sampling periods. The gestation months of the females were recorded for five trimesters by ultrasonography, as well as births and pregnancy losses. Recorded seropositivity and positivity for agent DNA were 90.9% (95% CI 58.7-9.7) and 36.3% (95% CI 10.9-69.2), respectively, on day zero. N. caninum DNA was detected between 18.1% (95% CI 2.3-51.7) and 45.4% (95% CI 16.7-76.6) over the five trimesters of observation, with three births and three abortions recorded. The studied water buffalo population had a high presence of N. caninum antibodies; however, the detection of N. caninum DNA remained below 47% in the females. The association was only observed in the detection of DNA with pregnant females (P 0.007). Our results support the hypothesis of the resistance of water buffaloes to infection and the onset of clinical signs against infection by N. caninum even upon a high possibility of infection and reinfection described in this production system in Mexico.

Detection of Bartonella bovis DNA in blood samples from a veterinarian in Mexico.

The genus Bartonella encompasses 38 validated species of Gram-negative, facultative intracellular bacteria that colonize the endothelial cells and erythrocytes of a wide spectrum of mammals. To date, 12 Bartonella species have been recorded infecting humans, causing diseases of long historical characterization, such as cat scratch fever and trench fever, and emerging bartonellosis that mainly affect animal health professionals. For this reason, this study aimed to report a documented case of Bartonella bovis infecting a veterinarian from Mexico by the amplification, sequencing and phylogenetic reconstruction of the citrate synthase (gltA) and the RNA polymerase beta-subunit (rpoB) genes, and to report the natural course of this infection. To our knowledge, this work is the first to report the transmission of B. bovis via needlestick transmission to animal health workers in Latin America.

Molecular detection of hemotropic Mycoplasma and Bartonella species in lice from sheep and goats of Mexico.

The knowledge of lice associated with small ruminants, especially sheep and goats, is scarce. In Mexico, there are historical reports of six species of chewing and sucking lice associated with Capra hircus and Ovis canadensis. However, the reports did not analyze the ecology of the infestations or the presence of potentially pathogenic bacteria. For this reason, the objectives of this study were i) to identify the species of lice associated with sheep and goats in three states of the Mexican Republic, ii) to characterize the infestations, and iii) to identify the presence of bacterial pathogens. From October 2019 to August 2021, six ranches with sheep and goats were sampled in the states of Hidalgo and Veracruz. Hosts were visually inspected, and lice were retrieved with forceps. The specimens were sexed and identified using morphological taxonomic keys. DNA extraction was performed individually, and a fragment of the cytochrome oxidase subunit 1 gene (COI) was amplified for the molecular identification of the specimens. Subsequently, Anaplasma, Bartonella, Ehrlichia, Mycoplasma, and Rickettsia were molecularly detected. Additionally, the infestations were characterized by calculating the prevalence and mean abundances. We collected 563 specimens of three species, Bovicola caprae, Bovicola ovis, and Linognathus africanus. The highest infestation levels were recorded for B. ovis (66.7%; 4.4) from Veracruz. Additionally, two Bartonella species were detected: Bartonella mellophagi in B. ovis and Bartonella capreoli in L. africanus. In contrast, Mycoplasma ovis was detected exclusively in one pool of B. ovis. This study provides new bacterial-ectoparasite associations and highlights the possible role of these neglected ectoparasites as vectors in the populations of sheep and goats from Mexico.

Confirmation of the presence of Rickettsia felis infecting Ornithodoros puertoricensis in Mexico.

Soft ticks are neglected competent vectors of a wide range of pathogenic microorganisms, among which bacteria of the genera Rickettsia and Borrelia stand out. In Mexico, previous studies have shown the presence of a member of the Ornithodoros talaje complex in the Virginia opossum (Didelphis virginiana Didelphimorphia: Didelphidae Kerr) from southeastern Mexico. However, its specific identification has not been achieved. Two D. virginiana were treated in a private clinic during the period of April-May 2022. Tick larvae were manually removed, DNA extraction was performed, and some genes from various bacterial and parasitic pathogens were amplified and sequenced. A total of 96 larvae were recovered, which were morphologically identified as Ornithodoros puertoricensis (Ixodida: Argasidae Fox); the 16 S sequences showed a similarity of 96.79%-99.51% with sequences of O. puertoricensis from Panama and Colombia. The presence of Rickettsia felis (Rickettsiales: Rickettsiaceae Bouyer et al.) was detected in 15 specimens from one host. The soft tick O. puertoricensis is recorded for the first time as an ectoparasite of the Virginia opossum in America and represents the second report for this soft tick in Mexico since 1963. This represents the most northern record of this tick species in its geographic distribution and brings a new soft tick-Rickettsia association.

Detection of Bartonella bovis DNA in blood samples from a veterinarian in Mexico

ABSTRACT The genus Bartonella encompasses 38 validated species of Gram-negative, facultative intracellular bacteria that colonize the endothelial cells and erythrocytes of a wide spectrum of mammals. To date, 12 Bartonella species have been recorded infecting humans, causing diseases of long historical characterization, such as cat scratch fever and trench fever, and emerging bartonellosis that mainly affect animal health professionals. For this reason, this study aimed to report a documented case of Bartonella bovis infecting a veterinarian from Mexico by the amplification, sequencing and phylogenetic reconstruction of the citrate synthase (gltA) and the RNA polymerase beta-subunit (rpoB) genes, and to report the natural course of this infection. To our knowledge, this work is the first to report the transmission of B. bovis via needlestick transmission to animal health workers in Latin America.

Molecular detection of Coxiella-like endosymbionts and absence of Coxiella burnetii in Amblyomma mixtum from Veracruz, Mexico.

Ticks are obligate ectoparasites associated with a wide range of vertebrate hosts, including domestic animals. Moreover, ticks are capable of transmitting many pathogens such as Coxiella. To date, Coxiella burnetii, the etiological agent of coxiellosis or Q fever, is the only valid species of the genera. Nevertheless, a wide range of agents denominated Coxiella-like have been detected in recent studies, mainly associated with ticks. The pathogenicity of these Coxiella-like agents is controversial as some of them can infect both birds and humans. In Mexico, knowledge about Q fever is scarce and limited to historical serological records, and there is an overall lack of molecular proof of any agent of the genus Coxiella circulating in the country. Therefore, the aim of this study was to detect the presence of Coxiella in ticks associated with cattle in all 10 regions of Veracruz, Mexico. To accomplish this objective, first, we identified ticks collected from cattle and horses in Veracruz. Then, for Coxiella detection, DNA extraction from ticks and PCR amplification of the 16S-rDNA of Coxiella was performed. Finally, we performed a phylogenetic reconstruction to determine the Coxiella lineages detected. From the 10 regions sampled we collected 888 ticks grouped in 180 pools, and only five Amblyomma mixtum from the locality of Castán, and one from Los Angeles from Tuxpan were found positive, which represents a frequency of 20% for each locality. This study represents the first attempt at molecular detection of Coxiella in ticks associated with cattle in the state of Veracruz, the major livestock producer in the country. The findings of the present study are relevant as they establish a precedent regarding the circulation of Coxiella-like agents, as well as the absence in three municipalities of the state of Veracruz of C. burnetii, an abortive agent of livestock importance.

Prevalence and risk factors of coccidiosis in calves from Veracruz, México.

The objectives of the present study were: (1) to determine the prevalence of Eimeria spp. sporulated oocysts in calves from 26 Municipalities in the Central Zone of the State of Veracruz, Mexico, (2) to identify the Eimeria spp. infecting calves, and (3) to identify the risk factors associated with the presence of Eimeria spp. in tropical cattle. A total of 930 individual fecal samples were analyzed by using the McMaster technique; then, oocysts were maintained in 2.5% potassium dichromate to allow sporulation. The general prevalence of calves with Eimeria spp. oocysts was 39.7% (370/930). Of a total of 10 identified species, Eimeria canadensis was the most observed, followed by Eimeria bovis and Eimeria zuernii. The statistical analysis showed an association between the age of the calves (5 to 9 months), the presence of other animals, the physiographic regions and the restricted type of husbandry with the presence of Eimeria spp. in calves (P<0.05). Protective risk factors, such as: routine coprological analysis was also associated with a decrease in infection. These data demonstrate the presence of coccidia in cattle from the State of Veracruz, additionally to the necessary measures that must be taken to control this parasitosis.

First report of Babesia bigemina in Lama glama in a zoological garden of Veracruz, Mexico.

Zoological gardens represent specialised centres for the preservation of biological inventories and genetic diversity, allowing the recognition of multiple species in critical conservation categories. However, the close coexistence of multiple species of vertebrates that may be associated with various species of ectoparasites may be the cause of the transmission of multiple infectious agents, among which tick-borne pathogens stand out. In these areas, several animal species usually live in a small space and proximity to other wildlife, visitors and keepers. In Mexico, little is known about the disease agents transmitted by arthropods in zoological gardens. For this reason, the aim of this study was to identify the presence of Babesia/Theileria in animals maintained in captivity. As a part of a project identifying vector-borne pathogens in wildlife, 24 animals were sampled in the Miguel Angel de Quevedo zoo. Molecular identification of Babesia/Theileria was realised through amplification of a fragment of the mitochondrial cytB gene and the ribosomal 18S-rDNA. Two neotropical camelids (Lama glama) tested positive (2/3 = 66.6%) to Babesia bigemina. Our results represent the first record of B. bigemina in animals in captivity in a zoological garden in Mexico and the first finding of this haemoparasite in neotropical camelids in Mexico.

No molecular evidence of SARS-CoV-2 infection in companion animals from Veracruz, Mexico.

Active epidemiological surveillance of infectious agents represents a fundamental tool for understanding the transmission dynamics of pathogens and establishing public policies that can reduce or limit their expansion. Epidemiological surveillance of emerging agents, such as the recently recognized severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the cause of COVID-19, is essential to establish the risk of transmission between species. Recent studies reveal that companion animals are organisms susceptible to being infected by this pathogen due to the close contact they have with their owners. For this reason, the aim of the present work was to detect the presence of SARS-CoV-2 in dogs and cats in the state of Veracruz, Mexico, where there is active transmission of this microorganism in human populations. Oral and nasopharyngeal swab samples were collected from dogs and cats with a history of exposure to patients with COVID-19. Total RNA was extracted and detection of viral genes N1 and N2 was performed by reverse transcription polymerase chain reaction (RT-qPCR). All 130 samples of companion animals tested by RT-qPCR for SARS-CoV-2 were negative at the time they were collected. This study represents the second active surveillance of SARS-CoV-2 in populations of domestic dogs and cats in Latin America and the first approach in Mexico. Given that coronaviruses have shown a high capacity to be transmitted between species, it is imperative to establish measures to prevent this agent from entering and establishing in populations of companion animals.

Spatial epidemiology of Leptospira sp. exposure in bovines from Veracruz, México.

Bovine leptospirosis is a bacterial disease that affects cattle herds, causing economic losses due to reproductive problems which require expensive treatments. The main source of transmission for cattle is still uncertain, but rodents and bats can play an important role in the transmission cycle by being maintenance hosts for the pathogenic species of the bacterium and spreading it through urine. In this study, we characterize possible risk areas for bovine leptospirosis exposure in the state of Veracruz, Mexico, based on the geographical distribution of flying (bats) and terrestrial (rodents and opossums) wild hosts of Leptospira sp. reported in Mexico, in addition to climate, geography, soil characteristics, land use and human activities (environmental variables). We used a generalized linear regression model to understand the association between the frequency of anti-Leptospira sp. antibodies (a proxy of exposure) in cattle herds exposed to Leptospira, the favourability of wild hosts of Leptospira as well as the environmental variables. The parameterized model explained 12.3% of the variance. The frequency of anti-Leptospira sp. antibodies exposure in cattle herds was associated with elevation, geographic longitude, pH of the soil surface and environmental favourability for the presence of rodents, opossums and bats. The variation in exposure was mainly explained by a longitudinal gradient (6.4% of the variance) and the favourability-based indices for wild hosts (9.6% of the variance). Describing the possible risks for exposure to Leptospira in an important and neglected livestock geographical region, we provide valuable information for the selection of areas for diagnosis and prevention of this relevant disease.

Molecular evidence of Leishmania spp. in spider monkeys (Ateles geoffroyi) from The Tuxtlas Biosphere Reserve, Veracruz, Mexico.

The black-handed spider monkey (Ateles geoffroyi) is a platyrrhine primate distributed in southern Mexico, Central America, and part of South America. Two subspecies inhabit Mexico: Ateles geoffroyi vellerosus and Ateles geoffroyi yucatanensis, both threatened with extinction. Serological evidence of exposure of spider monkeys to various groups of parasites such as Trypanosoma cruzi in México and Leishmania spp. in Brazil has been reported. The genus Leishmania encompasses about 23 species of flagellate protozoa that are transmitted by the bite of females of Phlebotominae sand flies. These parasites cause a zoonotic disease called leishmaniasis, which generates skin, mucocutaneous and/or visceral manifestations. The aim of the present study was to demonstrate the presence of Leishmania sp. in spider monkeys from the Tuxtlas Biosphere Reserve, Veracruz, Mexico. Blood samples from 10 free- ranging specimens of A. geoffroyi yucatanensis and 11 specimens in captivity of A. geoffroyi vellerosus were collected and used. The samples were subjected to a conventional Polymerase Chain Reaction test for the identification of a 116 bp fragment of a region from the kinetoplast minicircle of the parasite. Our analyzes showed that 71.4% of the sampled animals had fragment sizes compatible with Leishmania spp. The implications involve the survival of the specimens and the possibility that these primates act as sentinels of the disease. Furthermore, it is the first report suggesting the presence of Leishmania spp. in A. geoffroyi vellerosus and A. geoffroyi yucatanensis in Veracruz, Mexico.

Serosurveillance and risk factors for Neospora caninum infection in water buffaloes (B. bubalis) from central and southern Mexico.

The water buffalo (B. bubalis) is an alternative to cattle ranching in several regions of southern Mexico. Here we report seroprevalence and risk factors associated with the protozoan parasite, Neospora caninum, in water buffaloes in six buffalo production units, in municipalities from central and southern Veracruz, Mexico. Antibodies to N. caninum were assessed in serum samples of 543 buffaloes by a commercial ELISA-kit, and 44.8% (243/543; 95% CI 40.5-49.0) were seropositive. Questionnaires were used to collect epidemiological data and to identify risk factors associated with N. caninum infection. Data analysis indicated that older buffaloes (≥7 year) exhibited the highest seroprevalence for neosporosis 62.3% (38/61; 95% CI 49.7-73.4) (P ≤ 0.05). Buffaloes that were in close contact with cattle had higher seroprevalence 47.6% (168/353; 95% CI 42.3-52.9) (P < 0.01) than those that were not in contact 36.8% (70/190; 95% CI 30.0-44.1). Our findings provide important information to implement preventive measures in the buffalo farms.

Morphometrics of Amblyomma mixtum in the State of Veracruz, Mexico.

The tick Amblyomma mixtum is an ectoparasite of veterinary and public health importance because of its role as a vector of zoonotic pathogens. However, little is known about A. mixtum intraspecific variability and if morphological differentiation exists between populations across its geographic range. This study aimed to determine by electron microscopy the morphological variability of A. mixtum populations in the state of Veracruz, which has a large livestock population among states in Mexico. Forty male and 40 female A. mixtum collected from the 10 natural regions of Veracruz state were analyzed microscopically to accomplish main component analysis for each sex. Clusters were calculated with the out-tree method and a dendrogram produced to group the specimens according to their morphometric characteristics. Using 10 main components, 77% of the morphological variation of the ticks was explained. This is a reflection of scarce intraspecific variation between females. The dendogram for females grouped six clusters of specimens with similar characteristics. Morphometric variability in males was described using eight main components. Limited intraspecific variation was also observed between males. In males, the dendogram yielded six groups with similar morphometric characteristics. Morphometric analyses confirmed that the only species from the Amblyomma cajennense complex that are parasites to livestock in Veracruz state is A. mixtum. The eryxenous nature of A. mixtum combined with the frequent movement of livestock hosts may contribute to the apparent homogeneous phenotype of this tick species in Mexico.

Molecular detection of Theileria equi in horses from Veracruz, Mexico.

Equine piroplasmosis is a tropical parasitic disease caused by several intraerythrocytic members of the genera Babesia and Theileria. These pathogens are transmitted by multiple tick species and are considered as important exotic microorganisms in several countries. Equine piroplasmosis causes significant economic losses due to abortions, decreased activity and even death of equines, making surveillance of these infectious disease essential. In the northern and eastern parts of Mexico, few molecular and serological reports have been made on the presence and exposure of horse to these agents. For this reason, the aim of the current work was to perform a molecular detection study of Babesia and Theileria in equines from the state of Veracruz, Mexico. A total of 100 whole blood samples were tested. Chelex-100 resin was used for DNA extraction and a fragment of 459 bp of the 18S rRNA gene of members of the genera Babesia/Theileria were identified. Of the 100 samples analysed, 18 tested positive for Babesia/Theileria, resulting in a prevalence of 18 %. Identity analyses and phylogenetic reconstruction revealed that all samples were infected with Theileria equi. This work represents the first molecular record of Babesia/Theileria in equines from the state of Veracruz, Mexico, and demonstrates the endemicity of T. equi in this region of the country.

The genus Rickettsia in Mexico: Current knowledge and perspectives.

The genus Rickettsia encompasses 35 valid species of intracellular, coccobacilli bacteria that can infect several eukaryotic taxa, causing multiple emerging and re-emerging diseases worldwide. This work aimed to gather and summarise the current knowledge about the genus Rickettsia in Mexico, updating the taxonomy of the bacteria and their hosts by including all the records available until 2020, to elucidate host-parasite relationships and determine the geographical distribution of each Rickettsia species present in the country. Until now, 14 species of Rickettsia belonging to four groups have been recorded in Mexico. These species have been associated with 26 arthropod species (14 hard ticks, three soft ticks, two sucking lice, and seven fleas) and 17 mammal species distributed over 30 states in Mexico. This work highlights the high biological inventory of rickettsias for Mexico and reinforces the need to approach the study of this group from a One Health perspective.

Integrative taxonomic description of the chewing louse Tricholipeurus lipeuroides infesting Odocoileus virginianus veraecrucis white-tailed deer in Veracruz, Mexico.

Chewing lice (Phthiraptera: Amblycera, Ischnocera) represent a component of the ectoparasite fauna associated with large sized mammals as deers. However, the diversity of chewing louse species infesting deer remains to be fully characterized in the Neotropics. Little is known about the chewing lice infesting the extant fourteen subspecies of white-tailed deer (Odocoileus virginianus) in Mexico. Known to infest white-tailed deer (WTD) in Canada and the United States (U.S.), Tricholipeurus lipeuroides is a chewing louse species that was originally described in the nineteenth century infesting O. v. mexicanus in Mexico. For the first time, infestation of O. v. veraecrucis, a Neotropical WTD subspecies in Mexico, with T. lipeuroides is reported herein. An integrative taxonomic approach was taken by combining morphological and molecular analyses to describe the T. lipeuroides infestion of O. v. veraecrucis. Ecological parameters of the T. lipeuroides infestations were also calculated. The prevalence was 91.7% of the 56 O. v. veraecrucis (29 females and 27 males) inspected while under chemical restraint that were sampled at 3 sites in the central region of Veracruz state in Mexico. The amplification and sequencing of previously reported T. lipeuroides Cytochrome Oxidase Subunit I gene confirmed the identity of all the chewing louse life stages. These results are discussed in the context of comparative analyses on the emergence of novel chewing lice-deer associations.

First record of natural infection with Anaplasma marginale in sucking lice infesting the water buffalo (Bubalus bubalis) in Mexico.

The water buffalo (Bubalus bubalis) is an Asian species of bovine which was introduced in Mexico in 1992, as an alternative for milk and meat production. However, little is known about its associated ectoparasites, in particular about their lice. As a part of a project to identify the diversity and prevalence of Anaplasma in cattle and water buffaloes in the state of Veracruz, Mexico, we visited a ranch in the downtown area in which 42 buffaloes were visually inspected for the presence of ectoparasites. Lice were manually recovered, and morphologically identified using taxonomic keys. They were also molecularly identified by the amplification of a fragment of the cytochrome oxidase C subunit I gene (COX1). As a result, we recovered 20 lice from eight animals. All lice were identified as Haematopinus tuberculatus, and the recovered sequences exhibited a similarity of 99.7% with those of the same species deposited in GenBank. Additionally, we detected the presence of Anaplasma marginale in 70% (14/20) of the analysed samples. Our results represent the first record of this exotic ectoparasite for the country.

SEROLOGIC SURVEY OF LEPTOSPIRA SPP. IN CAPTIVE ANIMALS FROM VERACRUZ, MEXICO.

Leptospirosis is the most common zoonotic disease worldwide and is considered endemic in countries with tropical climates. It is caused by 10 species of the Leptospira genus and by more than 275 serovars which can affect a wide range of vertebrates. In the Americas, 122 species of four classes of vertebrates have been reported to be infected or exposed to many Leptospira species. Many of these reports are from zoos and rehabilitation centers. Mexico has one single study that reported antibody titers against Leptospira in zoo animals. The purpose of this research was to identify the degree of exposure of some captive mammals and reptiles in Veracruz, a Mexican state with endemic leptospirosis, through microagglutination using 14 live strains of five Leptospira species. Sera samples were collected from 55 animals of 11 species from two classes (Mammalia and Reptilia), four orders (Primates, Artiodactyla, Carnivora, Crocodilia), and nine genera. The more prevalent serovars were Icterohaemorrhagiae and Tarassovi and the highest titers were reactive to the serovar Icterohaemorrhagiae with a value of 1: 51,200.

Molecular detection of reptile-associated Borrelia in Boa constrictor (Squamata: Boidae) from Veracruz, Mexico.

The reptile-associated Borrelia represent a monophyletic group of bacteria transmitted by several species of hard ticks, which has been reported to only infect amphibians and reptiles in Eurasia and Middle East, however, this bacterial group has not been studied in North America. The aim of this study was to assess the presence of Borrelia spirochetes in blood samples of native reptiles of Mexico. Blood samples were directly obtained from individuals, DNA extractions were performed using Chelex-100. The Borrelia detection was performed by conventional PCR. From 102 reptiles tested, only five individuals of Boa constrictor were positive for the presence of DNA of the reptile-associated Borrelia group. Supported by phylogenetic analysis, this study presents the first record of these spirochetes group in Mexico, and initial evidence of B. constrictor as a host of this group.